Cytokines comprise a large family of secreted proteins that regulate cell growth and differentiation of many types of cells. These factors are especially important in regulating immune and inflammatory responses, regulating lymphoid development and differentiation. Cytokines also regulate immune homeostasis, tolerance, and memory. Not surprisingly, cytokines are critical in the pathogenesis of autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease and psoriasis. Understanding the molecular basis of cytokine action provides important insights into the pathogenesis of immune-mediated disease and offers new therapeutic targets. We discovered haman Jak3, a kinase essential for signaling by cytokines that bind the common gamma chain, gc (IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21). We found that mutation of Jak3 results in a primary immunodeficiency disorder termed severe combined immunodeficiency (SCID). We have received two patents related to targeting Jak3 as the basis for a new class of immunosuppressant/immunomodulatory drugs and established a CRADA with Pfizer to generate a the first generation Jak3 antagonists. One compound, tofacitinib (CP 690,550), was produced by Pfizer and found to be effective in preclinical models. The drug is being tested presently in Phase III studies in rheumatoid arthritis, psoriasis and transplant rejection, where it appears to be efficacious. The CRADA with Pfizer was renewed this year and was directed at better understanding the mechanisms of action of this drug. We began by assessing the effect of tofacitinib on biochemical pathways activated by IL-2. There has been debate over the years regarding Jak-dependent and independent aspects of cytokine signaling. However, to date, we have found that all substrates phosphorylated in response to IL-2 are abrogated by tofacitinib. These include MAPK and AKT activation, as well as STAT activation. We have also determined that tofacitinib has dramatic effects on cytokine-dependent T cell proliferation and viability. We next assessed the effect of ttofacitinib on CD4+ helper cell differentiation. CD4+ T cells have a number of potential fates. In addition to the well-known helper T cell fates, T helper 1 and T helper 2 cells, other fates are now recognized. A more recently recognized subset of CD4+ T cells is a subset that preferentially produce the cytokine IL-17 (Th17 cells). IL-17 is a major inflammatory cytokine, which appears to contribute to the pathogenesis of many autoimmune and autoinflammatory disorders including rheumatoid arthritis, spondyloarthropathy, multiple sclerosis and inflammatory bowel disease. We found that tofacitinib blocked Th1 and Th2 differentiation. The latter can be explained by effects on IL-4 signaling, which is dependent upon gc/Jak3. We found that the former was also blocked and this was because tofacitinib blocked IFNg signaling via effects on Jak1 and also inihibited IL-12, another key driver of Th1 differentiation. Tofacitinib was also found to block IL-23-dependent generation of Th17 cells. The effects on tofacitinib on in vivo models of autoimmunity were also examined. We found that the Jak inhibitor is very effective in attenuating disease in arthritis models. Importantly, we have also found that tofacitinib has actions beyond effects on gc cytokines. Specifically, we investigated the effect of this drug on innate immune responses and found that the drug blocks inflammation in an acute sepsis model. We believe that these effects represent important aspects of the efficacy of this drug in rheumatoid arthritis and other autoimmune disorders. Current studies are ongoing evaluating tofacitinib in models of graft versus host disease. In addition, we are studying the effects of tofacitinib on T cell metabolism. In more recent studies done in collaboration with NCI, we have investigated the efficacy of tofacitinib as an adjunct in immunotoxin therapy for the treatment of cancer. In a mouse model, we have found that tofacitinib very effectively blocks immune responses to immunotoxins. This is the subject of a recently submitted patent.